The regulation of growth in vertebrates is believed to involve the interaction of the mechanisms for the release of growth hormone with the 14-amino acid peptide somatostatin or its various alternative forms. At least one function of somatostatin in vivo is to depress growth, presumably by interaction with this system.
Accordingly, it has been suggested that antibodies reactive with somatostatin should be capable of stimulating growth by depressing the action of somatostatin with the growth regulatory system, and it has been shown that rats administered antisomatostatin antiserum show increases in growth hormone levels; in particular the decrease in growth hormone levels ordinarily observed in response to stress is mitigated (Chihara, K., et al, Endocrin (1978) 103:1916; Arimura, et al, Endocrin (1976) 98:540).
Accordingly, attempts have been made to depress somatostatin activity using the immune system. Spencer, et al (Animal Production (1981) 31:376; Veterinary Record (May 22, 1984), p. 484) have shown that lambs injected with somatostatin linked to carrier protein raised antibodies against the conjugate and showed enhanced weight gain. Varner et al, Endocrinology (1980) 106:1027, reported that growth hormone concentrations in serum were higher in lambs autoimmunized against somatostatin than in controls; however, growth was not enhanced in these animals. Additional reports of attempts to use this system to enhance growth or demonstrate increases in levels of growth hormone include Lovinger, R., et al, Endocrinol (1974) 95:743; Kato, Y., Endocrinol (1974) 95:1608.
The interaction of somatostatin with the growth hormone system has also been demonstrated by regulating somatostatin levels directly. Cowan, J. S., Canadian J Physiol Pharmacol (1984) 62:199-207, showed withdrawal of somatostatin initiated bursts of growth hormone secretion in dogs. It has also been shown that growth hormone releasing factor (GRF) will result in elevated growth hormone concentration only at low somatostatin levels (Cowan, J. S., et al, Canadian J Physiol Pharmacol (1985) 63:AIX (Abstract)).
The production of monoclonal antibodies which are immunoreactive with somatostatin has been reported. Buchan, A. M. J., et al, Histochemistry (1985) 83:175-180, describe the production of antibodies initially screened for immunoreactivity with somatostatin. To produce these antibodies, somatostatin-14 (cyclic) was conjugated to keyhole limpet hemocyanin (KLH) (Cal Biochem) using carbodiimide and dialyzed overnight at 4.degree. C. The dialyzed preparation was used to immunize BIO.BRSgSn mice (Jackson Laboratories, Bar Harbor, Me.) which were injected three times with 30 nM of the conjugate, and the sera were titrated for antisomatostatin using an ELISA assay. The spleen from the mouse showing the best antibody response was fused with NSI cells according to the method of Kohler and Milstein, Eur J Immunol (1976) 6:511-521, as modified by Fazekas de St. Groth, et al, J Immunol Meth (1980) 35:1-21, and of Oi, V. T., et al, Selected Methods in Cellular Immunology (1980), Michel, B. B., et al, eds. W. H. Freeman, San Francisco.
Clones were screened using the ELISA method of Boller, A., et al, Bull WHO (1976) 53:55-65. Positively testing clones were grown and the supernatants retested for specificity in ELISA against somatostatin, KLH, and an unrelated antigen, ferredoxin.
Four hybrids which showed the correct specificity for somatostatin were cloned out by limiting dilution 5 times and grown as ascites tumors of irradiated outbred mice. The antibodies could be partially purified from the fluid by precipitation by 50% ammonium sulfate, and then dialyzed and lyophilized.
These four antibodies were shown to be immunocytochemically reactive with a number of cells associated with the neural and digestive system. A description of the histochemical activity of these antibodies is set forth in the above-referenced article by Buchan et al and in the report by Vincent, S. R., et al, J Compar Neurol (1985) 238:169-186.
A U.S. Pat. No. 4,599,229, claims methods to promote growth using both polyclonal antisera against somatostatin and monoclonal antibodies prepared from hybridomas formed by fusions involving lymphocytes from immunized animals. The disclosure proposes screening hybridoma supernatants against somatostatin per se, however, and fails to disclose that only a fraction of antibodies thus screened will in fact be effective in enhancing growth.